CN111411057B - Lactobacillus corynebacterium for producing broad-spectrum antibacterial peptide and application of antibacterial peptide thereof - Google Patents

Lactobacillus corynebacterium for producing broad-spectrum antibacterial peptide and application of antibacterial peptide thereof Download PDF

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CN111411057B
CN111411057B CN202010319752.XA CN202010319752A CN111411057B CN 111411057 B CN111411057 B CN 111411057B CN 202010319752 A CN202010319752 A CN 202010319752A CN 111411057 B CN111411057 B CN 111411057B
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汪少芸
韩金志
孟晓洁
沈昊
姚思羽
陈旭
蔡茜茜
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Abstract

The invention provides a lactobacillus corynebacterium for producing broad-spectrum antibacterial peptide and application of the antibacterial peptide thereof, belonging to the technical field of microorganisms. Lactobacillus corynebacterium of the invention: (Lactobacillus coryniformis) FZU63, having a accession number: CGMCC NO. 19553. The antibacterial peptide produced by the strain has obvious inhibiting effect on various gram-positive bacteria and gram-negative bacteria. The antibacterial peptide AMP-fzu63 produced by Lactobacillus coryniformis FZU63 comprises 2 polypeptide sequences having the sequence RQQPMTLDYRW, LVKNAKERGY. The antibacterial peptide AMP-fzu63 generated by the lactobacillus corynebacterium FZU63 is applied to the fresh-keeping of marine fish, and can reduce the generation of unpleasant odor substances hexanal and 1-octen-3-ol by inhibiting the growth and reproduction of microorganisms in the fish.

Description

Lactobacillus corynebacterium for producing broad-spectrum antibacterial peptide and application of antibacterial peptide thereof
Technical Field
The invention belongs to the field of microorganisms, and particularly relates to a lactobacillus corynebacterium for producing broad-spectrum antibacterial peptide and application of the antibacterial peptide.
Background
Putrefaction and deterioration of fish meat are generally caused by three factors, namely autolytic enzyme decomposition, fat oxidation and microbial breeding; among them, microbial growth and metabolism are the main causes of putrefaction of fish meat. After various seawater fishes are produced in coastal areas of China, most of the seawater fishes are difficult to survive due to sudden changes of environmental factors such as air pressure, temperature, dissolved oxygen and the like after being caught and leave water. After the marine fish dies, cell metabolism is terminated, and epidermal goblet cells stop secreting mucus containing antibacterial substances; in addition, the fish meat is short in muscle fiber and rich in moisture, so that the fish meat is easy to decay and deteriorate due to microbial breeding in the storage process, and huge loss is caused. Studies have shown that Shewanella, Pseudomonas, Aeromonas, Vibrio, Photorhabdus, Bacillus, and Enterobacter partially, etc., are specific putrefying bacteria (SSOs) in marine fish, and play a dominant role in the putrefying process of marine fish. Meanwhile, part of food-borne pathogenic bacteria pollute the marine fishes, and particularly Listeria monocytogenes, salmonella, hemolytic escherichia coli, vibrio parahemolyticus and the like bring serious threats to human health.
Lactic acid bacteria are GRAS (generally fermented As safety) grade microorganisms, and have been widely used for food fermentation and preservation. Numerous studies have shown that some lactic acid bacteria such as lactobacillus plantarum, lactobacillus acidophilus, lactobacillus casei, lactobacillus corynebacterium, lactobacillus delbrueckii, etc. produce extracellular bacteriocins (antibacterial peptides) that exhibit different bacteriostatic activities against gram-positive bacteria, gram-negative bacteria and fungi. The lactobacillus source bacteriocin has the characteristics of strong antibacterial activity, high safety and the like, and is a green biological preservative with wide application prospect. At present, a large number of researches on The separation and purification, structure identification and bacteriostatic activity of lactobacillus-derived bacteriocins such as Plantaricin, lactococcus, Acidocin and Lactocin are reported at home and abroad, and 43 lactobacillus-derived bacteriocins are collected in an antibacterial peptide online database (3 months 2020). However, most bacteriocins are directed against gram-positive bacteria (G)+Bacteria) and against gram-negative bacteria (G)–Bacteria) has limited antibacterial effect, and only a few bacteriocins such as lactobacillus plantarum can inhibit G–And (5) bacteria. Bacteriocins such as Plantaricin ST28MS and ST26MS produced by Lactobacillus plantarum inhibit Pseudomonas aeruginosa and Escherichia coli; the Plantaricin MG can inhibit salmonella typhimurium and escherichia coli; plantaricin BacTN635 can inhibit Salmonella enteritidis, Pseudomonas aeruginosa, Hafnia and Serratia. To date, only two bacteriocins of lactic acid bacteria origin, Nisin (Nisin) and Pediocin PA-1, have been approved for use as food preservatives, but inOnly Nisin is allowed to be applied to food in China. Nisin is only for G+The bacteria have antibacterial activity on G–The bacteria have no bacteriostatic effect, and are limited to G of Shewanella putrefaciens, Pseudomonas and the like–The application of the bacteria as special putrefying bacteria in the preservation of fresh marine fish. At present, the food safety is highly valued in China, and because of the food safety risk of the chemical synthesis preservative, the novel, green, safe and broad-spectrum lactobacillus bacteriocin is developed, so that the preservative has important significance for preventing and preserving marine fishes, inhibiting the breeding of microorganisms and inhibiting the pollution of food-borne pathogenic bacteria. The broad-spectrum antibacterial peptide derived from the lactic acid bacteria is a green biological preservative with wide application prospect due to the characteristics of strong antibacterial activity, wide antibacterial spectrum, high safety and the like.
Disclosure of Invention
The invention aims to provide a corynebacterium strain for producing broad-spectrum antibacterial peptide and application of the antibacterial peptide thereof aiming at the defects of the prior art.
In order to achieve the purpose, the invention adopts the following technical scheme:
a corynebacterium strain producing broad-spectrum antibacterial peptide, wherein the corynebacterium strain producing broad-spectrum antibacterial peptide is corynebacterium (L.) (Lactobacillus coryniformis) FZU63, which is classified and named: lactobacillus corynebacterium: (Lactobacillus coryniformis) And is preserved in China general microbiological culture Collection center (CGMCC) in 2020, 4 and 7 months, with the preservation number as follows: CGMCC number 19553, the preservation address is as follows: xilu No.1 Hospital No. 3, Beijing, Chaoyang, North.
The above-mentioned Lactobacillus corynebacterium (C.), (Lactobacillus coryniformis) FZU63 is obtained by separating Chinese cabbage from rural families in elm city of Jilin province.
Lactobacillus corynebacterium: (Lactobacillus coryniformis) FZU63 gram stain positive, the colony is round, whitish, small and convex, and the edge is neat.
Lactobacillus corynebacterium: (Lactobacillus coryniformis) FZU63 has broad-spectrum antibacterial activity, and can be used for treating Staphylococcus aureus ATCC 6538, Staphylococcus aureus ATCC 12600, and methicillin-resistant Staphylococcus aureusStaphylococcus MW2, methicillin-resistant Staphylococcus aureus N315, Listeria monocytogenes CICC 21633, Micrococcus luteus CMCC 28001, 6 gram-positive bacteria and Escherichia coli ATCC 25922, Escherichia coli CMCC (B) 44102, Escherichia coli enterohemorrhagic 21530, Vibrio parahaemolyticus ATCC 17802, Salmonella choleraesuis CICC 13312, Salmonella typhimurium CICC 51005, Salmonella enteritidis CICC 21527, Salmonella paratyphi A CMCC (B) 50093, Salmonella paratyphi B CMCC (B50094), Salmonella sakazakii ATCC 29544, Pseudomonas fluorescens AS1.55, Shewanella ATCC 138, 12 gram-negative bacteria all show good bacteriostatic effects.
The above-mentioned broad-spectrum antibacterial peptide-producing coryneform bacterium, i.e., Lactobacillus corynebacterium: (Lactobacillus coryniformis) FZU63, wherein the antimicrobial peptide AMP-3652 comprises 2 polypeptide sequences, and the specific polypeptide sequence is as follows: RQQPMTLDYRW, LVKNAKERGY are provided.
The application of the antibacterial peptide AMP-fzu63 in the fresh-keeping of the marine fish is provided.
The invention has the advantages that: the invention obtains a lactobacillus corynebacterium FZU63 strain producing broad-spectrum antibacterial peptide by screening, and the antibacterial peptide produced by lactobacillus corynebacterium FZU63 has obvious inhibition effect on various gram-positive bacteria and gram-negative bacteria. The antibacterial peptide AMP-fzu63 produced by Lactobacillus coryniformis FZU63 comprises 2 polypeptide sequences having a polypeptide sequence of RQQPMTLDYRW, LVKNAKERGY. The antibacterial peptide AMP-fzu63 generated by the lactobacillus corynebacterium FZU63 is applied to the fresh-keeping of the marine fish, can reduce the generation of unpleasant odor substances hexanal and 1-octen-3-ol by inhibiting the growth and reproduction of microorganisms in the marine fish, and has the effect obviously superior to that of the lactobacillus-derived antibacterial peptide Nisin (Nisin) used for food preservation and fresh-keeping in the prior art.
Description of the drawings:
FIG. 1 is a gel electrophoresis image of PCR amplification products. FZU63, FZU72, FZU73, FZU101, FZU105, FZU122 and Marker are sequentially arranged from left to right.
FIG. 2A phylogenetic tree of FZU63 strain was constructed based on the 16S rDNA gene sequence.
FIG. 3 shows the bacteriostatic effect of Lactobacillus corynebacterium FZU63 fermented liquid on gram-positive bacteria. Staphylococcus aureus ATCC 6538, B: staphylococcus aureus ATCC 12600, C: methicillin-resistant staphylococcus aureus MW2, D: methicillin-resistant staphylococcus aureus N315, E: listeria monocytogenes CICC 21633, F: micrococcus luteus CMCC 28001.
FIG. 4 shows the bacteriostatic effect of the fermentation broth of Lactobacillus corynebacterium FZU63 on gram-negative bacteria. A: escherichia coli ATCC 25922, B: escherichia coli CMCC (B) 44102, C: enterohemorrhagic escherichia coli cic 21530, D: vibrio parahaemolyticus ATCC 17802, E: salmonella choleraesuis cic 13312, F: salmonella typhimurium CICC 51005, G: salmonella enteritidis CICC 21527, H: salmonella paratyphi type a cmcc (b) 50093, I: salmonella paratyphi b cmcc (b) 50094, J: enterobacter sakazakii ATCC 29544, K: pseudomonas fluorescens AS1.55, L: shewanella ATCC 49138.
FIG. 5 shows the inhibiting effect of the fermentation liquid of Lactobacillus corynebacterium FZU63 on the fungus Trichoderma viride CGMCC 3.2942.
FIG. 6 Sephadex LH-20 gel chromatogram of the sample.
FIG. 7 is the reversed phase HPLC chromatogram of the crude extract of the fermentation liquid of Lactobacillus corynebacterium FZU63 and the bacteriostatic activity of different separated components.
FIG. 8 shows the bacteriostatic activity of the reversed phase HPLC chromatography of different fractions of crude extract of fermentation broth of Lactobacillus corynebacterium FZU 63. C in the figure: mobile phase (20% methanol); 1. 2, 3, 4, 5 are the collected products of elution peaks 1, 2, 3, 4, 5 in fig. 7.
FIG. 9 is a secondary mass spectrum of the amino acid sequence RQQPMTLDYRW polypeptide of antimicrobial peptide AMP-fzu 63.
FIG. 10 is a secondary mass spectrum of the amino acid sequence LVKNAKERGY polypeptide of antimicrobial peptide AMP-fzu 63.
FIG. 11 shows the effect of the antimicrobial peptide AMP-fzu63 in the preservation of sea bass in cold storage. A, influence of AMP-fzu63 on colony count of sea bass in the process of refrigeration and preservation; b, the influence of AMP-fzu63 on the hexanal content of the fish meat of the sea bass in the process of refrigeration and preservation; c, the influence of AMP-fzu63 on the content of 1-octen-3-ol in fish meat during the refrigeration and preservation of sea bass.
Detailed Description
In order to make the present invention more comprehensible, the technical solutions of the present invention are further described below with reference to specific embodiments, but the following examples are only examples of the present invention and do not represent the scope of the present invention defined by the claims.
Example 1
The lactobacillus corynebacterium FZU63 is obtained by separating the Chinese cabbage from the Chinese cabbage prepared in rural families in elm city of Jilin province.
The specific separation method comprises the following steps: taking 10 g of Chinese cabbage, performing aseptic crushing, using aseptic normal saline for resuspension, performing gradient dilution by 10 times, respectively coating the mixture on MRS solid medium plates, and performing static culture for 18 h at 37 ℃; selecting single bacterial colony, placing in MRS liquid culture medium, standing and culturing at 37 deg.C for 24 h, centrifuging to obtain supernatant, and screening bacterial strains producing antibacterial substances by solid agar punching bacteriostasis method with Staphylococcus aureus and Salmonella typhimurium as indicator bacteria under the condition of eliminating interference of organic acid, hydrogen peroxide and other substances.
The gram stain of the strain producing the antibacterial substance is positive, the bacterial colony is round, whitish, small and convex, the edge is neat, and the characteristics are the same as the characteristics of a typical lactobacillus bacterial colony. 6 strains of bacteria with the inhibition zone diameter of more than 10 mm are selected from the lactobacillus strains and named as FZU63, FZU72, FZU73, FZU101, FZU105 and FZU122 respectively, the 6 strains are identified through GB4789.35-2016, the identification results are shown in Table 1, and the lactobacillus strains are primarily determined.
TABLE 1 Biochemical reaction test results of primary screening of 6 strains
Figure 907646DEST_PATH_IMAGE001
Note: + indicates that the strain is positive.
FZU63 the strain is inoculated in MRS liquid culture medium, activated and transferred to be cultured to logarithmic growth phase at 37 ℃. Taking 1-2 mL of culture solution, centrifuging for 1 min at 10000 g/min, discarding supernatant, and extracting total DNA from somatic cells by using a commercial bacterial total DNA extraction kit according to an instruction. PCR amplification was performed using 16S rDNA gene primers 16SAF (5'-AGAGTTTGATCCTGGCTCAG-3') and 16SAR (5 '-TACGGYTACCTTGTTACGACTT-3'), and 5. mu.L of the PCR product was subjected to 1% agarose gel electrophoresis, and the results of the electrophoresis are shown in FIG. 1.
PCR products were recovered by purification with a kit, and 16S rRNA full sequence sequencing was performed by Biotechnology engineering (Shanghai) Ltd. The 16S rDNA sequence of the strain FZU63 is 1242bp in total, and the sequencing result is shown in SEQ ID NO. 1.
SEQIDNO.1：
CTTTGGCTATCACTTTTGGATGGTCCCGCGGCGTATTAGTTAGTTGGTAAGGTAACGGCTTACCAAGACAATGATACGTAGCCGACCTGAGAGGGTAATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGAAAGTCTGATGGAGCAACGCCGCGTGAGTGAAGAAGGGTTTAGGCTCGTAAAACTCTGTTGTTAGAGAAGAACACATATGAGAGTAACTGTTCATCTTTTGACGGTATTCAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTTTTAAGTCTGATGTGAAAGCCTTCGGCTTAACCGAAGAAGTGCATCGGAAACTGGGAAACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACCCTGGTAGTCCATACCGTAAACGATGAATGCTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGCATTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTTTGCCCATCGTAGAGATAAAACTTTCCCTTCGGGGACAAAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATTACCAGTTGCCAGCATTTAGTTGGGCACTCTGGTGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGACGGTACAACGAGTTGCGAACTCGCGAGGGTAAGCTAATCTCTTAAAGCCGATCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTTTGTAACACCCAAAGTCGGTGGGGTAACCT
The 16S rDNA sequence of FZU63 strain was aligned in NCBI (https:// www.ncbi.nlm.nih.gov /) using BLAST. The strain can be classified as lactobacillus corynebacteria by comprehensive physiological and biochemical identification and 16S rDNA homology comparison resultLactobacillus coryniformis) The phylogenetic tree was constructed as shown in FIG. 2.
Example 2
Inoculating Corynebacterium-shaped lactic acid bacteria FZU63 in MRS liquid culture medium, standing and culturing at 37 deg.C for 24 h, centrifuging to obtain supernatant, respectively using Staphylococcus aureus ATCC 6538, Staphylococcus aureus ATCC 12600, methicillin-resistant Staphylococcus aureus MW2, methicillin-resistant Staphylococcus aureus N315, Listeria monocytogenes CICC 21633, Micrococcus luteus CMCC 28001, Escherichia coli ATCC 25922, Escherichia coli CMCC (B) 44102, Escherichia coli CICC 21530, Vibrio parahaemolyticus ATCC 17802, Salmonella choleraesuis CICC 12, Salmonella typhimurium CICC 51005, Salmonella enteritidis CICC 21527, Salmonella paratyphi CMCC (B) 50093, Salmonella paratyphi CC CMB (B) 50094, Salmonella sakazakii ATCC 29544, Pseudomonas fluorescens AS1.55, Shewanella ATCC49138, Trichoderma viride CGMCC3.2942 AS indicator bacteria, bacteria are inoculated on LB solid agar, trichoderma viride is inoculated on PDA solid culture medium, and the antibacterial effect of the antibacterial agent on the indicator bacteria is evaluated by adopting a punching antibacterial method. As shown in Table 2, the results of the agar pore diffusion experiments show that the fermentation supernatant of Lactobacillus corynebacterium FZU63 has a broad-spectrum bacteriostatic activity, and shows good bacteriostatic effects on 18 strains of bacteria, including 6 gram-positive bacteria (see FIG. 3) and 12 gram-negative bacteria (see FIG. 4), and has no bacteriostatic effect on fungi (see FIG. 5).
TABLE 2 Lactobacillus corynebacterium FZU63 fermentation liquid diameter of inhibition zone for different indicator bacteria
Figure 933370DEST_PATH_IMAGE002
Example 3
The lactobacillus corynebacterium FZU63 is subjected to static anaerobic culture for 36 h under the condition of 37 ℃ in an MRS liquid culture medium, centrifuged to take the supernatant, added with 3 times of ethanol by volume, stood for 12 h at 4 ℃, centrifuged to take the supernatant, and concentrated and removed with ethanol by a vacuum condensation rotary evaporator. Adding 5 mL of the concentrated sample into a Sephadex LH-20 gel column, isocratically eluting with 20% methanol at a flow rate of 0.2 mL/min, collecting 2 mL of the concentrated sample per tube, and verifying whether each tube has antibacterial activity and activity, wherein the elution curve is shown in FIG. 6. Collecting the tube with antibacterial activity, concentrating with rotary evaporator, and storing at 4 deg.C.
Crude extract of fermentation supernatant of lactobacillus corynebacterium FZU63 is obtained by Sephadex LH-20 filtration chromatography, FIG. 7 is reversed phase HPLC chromatography of the crude extract, elution peaks (1, 2, 3, 4, 5) with different retention times are collected, concentration is carried out by a rotary evaporator, the bacteriostatic activity is measured by taking Salmonella typhimurium CICC21483 as indicator bacteria, and FIG. 8 is obtained, wherein peaks 1, 3 and 4 have bacteriostatic activity. The activity peak 4 with the retention time of 9.193 min has the strongest antibacterial activity, the diameter of the antibacterial zone reaches 18.82 mm, and the protein content is 4.426 mg/mL by using a micro nucleic acid protein determinator. And collecting the active peak to carry out next structural identification by high-resolution mass spectrometry.
Example 4
The separated components with bacteriostatic activity obtained by separation and purification of a High Performance Liquid Chromatography (HPLC) are further analyzed by a nanoliter quadrupole time of flight tandem mass spectrometry (NLM) to identify the molecular structure of the antimicrobial peptide AMP-fuz 63. As shown in Table 3, 2 polypeptide sequences were obtained by mass spectrometry, and no identical sequence was found by alignment of an online Antimicrobial peptide database (antibacterial peptide database: http:// APs. unmac. edu/AP/main. php). As a result, 2 novel antibacterial peptides derived from Lactobacillus corynebacterium were identified (the secondary mass spectrograms of the 2 antibacterial peptides are shown in FIGS. 9 and 10).
TABLE 3 amino acid sequence of the antimicrobial peptide AMP-fuz63 after mass spectrometric identification
Figure 652934DEST_PATH_IMAGE003
Example 5
Killing fresh sea bass, removing fish scales and internal organs, cutting, weighing, placing in an aseptic sealed bag, adding antimicrobial peptide AMP-fzu63 at a ratio of 128 μ g/g according to the weight of fish, mixing uniformly, sealing, storing at 4 deg.C, setting aseptic normal saline treatment and Nisin treatment control groups, and sampling in 3, 7, 11 and 15 days respectively.
(1) Mixing fish meat with normal saline at a ratio of 1:10, homogenizing, diluting in gradient, coating on LB solid culture medium, culturing at 37 deg.C for 18 h, and counting colonies.
(2) Weighing 3-4 g of fish, cutting, placing in a 12 mL headspace extraction bottle, adding 5 mL of ultrapure water, 1 g of NaCl, a magnetic rotor and 10 muL of 10 mg/L2-octanol (an internal standard substance), rapidly sealing the sample bottle by using a sealing gasket, placing on a magnetic stirring heating table, stirring at the speed of 600 r/min, incubating for 20 min in a water bath at 40 ℃, inserting an aged solid phase microextraction needle, ejecting the extraction fiber to be fixed at a position 0.5-1.0 cm away from the liquid level, stirring while adsorbing in the headspace, recovering the extraction fiber after extracting for 30 min, pulling out the extraction needle, and inserting into a gas chromatography-mass spectrometer for analysis. Before the extraction by using the extraction head for the first time, the extraction head is aged to have no impurity peak at 250 ℃, and then extraction is carried out, and volatile odor substances in the fish are measured by a gas chromatography-mass spectrometer. Gas chromatography conditions: the temperature of a sample inlet is 250 ℃; keeping the initial temperature of the column oven at 40 ℃ for 8 min, then heating to 150 ℃ at 4 ℃/min, then heating to 250 ℃ at 20 ℃/min, and keeping for 5 min; no-shunt sample introduction, carrier gas: 99.999% high purity helium, carrier gas flow rate: 1 mL/min. Mass spectrum conditions: the ion source temperature is 230 ℃, the ionization mode is EI, the ionization energy is 70 eV, the interface temperature is 250 ℃, the quadrupole rod temperature is 150 ℃, the SCAN mode is selected as the scanning mode to perform qualitative analysis, the scanning range of ion fragments is 30-500 m/z, and the solvent delay time is 2.5 min.
As shown in FIG. 11A, the total number of colonies in the fish flesh of the sea bass treated with the antimicrobial peptide AMP-fzu63 was significantly lower than in the normal saline and Nisin treated groups as the refrigeration time was extended. The AMP-fzu63 can effectively inhibit the growth of microorganisms in fish meat and delay the putrefaction and deterioration of the sea bass caused by the growth and reproduction of the microorganisms.
As shown in FIGS. 11B and 11C, the contents of hexanal and 1-octen-3-ol, which are unpleasant odor substances, in fish meat during the storage period of sea bass showed a tendency of increasing and then decreasing with the lapse of the refrigerating time, and reached a maximum value at 7 to 9 days. In the antimicrobial peptide AMP-fzu 63-treated group, the contents of hexanal and 1-octen-3-ol in fish meat were significantly lower than those in the normal saline and Nisin-treated group with prolonged refrigeration time. Studies have shown that the changes of hexanal and 1-octen-3-ol content in fish meat have a close correlation with the growth activity of microorganisms. Therefore, the antibacterial peptide AMP-fzu63 can reduce the generation of the unpleasant odor substances hexanal and 1-octen-3-ol by inhibiting the growth and reproduction of microorganisms in the fish meat of the sea bass.
The above description is only a preferred embodiment of the present invention, and all equivalent changes and modifications made in accordance with the claims of the present invention should be covered by the present invention.
SEQUENCE LISTING
<110> Fuzhou university
<120> lactobacillus corynebacterium strain for producing broad-spectrum antibacterial peptide and application of antibacterial peptide thereof
<130> 5
<160> 5
<170> PatentIn version 3.3
<210> 1
<211> 1242
<212> DNA
<213> Artificial sequence
<400> 1
ctttggctat cacttttgga tggtcccgcg gcgtattagt tagttggtaa ggtaacggct 60
taccaagaca atgatacgta gccgacctga gagggtaatc ggccacattg ggactgagac 120
acggcccaaa ctcctacggg aggcagcagt agggaatctt ccacaatgga cgaaagtctg 180
atggagcaac gccgcgtgag tgaagaaggg tttaggctcg taaaactctg ttgttagaga 240
agaacacata tgagagtaac tgttcatctt ttgacggtat tcaaccagaa agccacggct 300
aactacgtgc cagcagccgc ggtaatacgt aggtggcaag cgttgtccgg atttattggg 360
cgtaaagcga gcgcaggcgg ttttttaagt ctgatgtgaa agccttcggc ttaaccgaag 420
aagtgcatcg gaaactggga aacttgagtg cagaagagga cagtggaact ccatgtgtag 480
cggtgaaatg cgtagatata tggaagaaca ccagtggcga aggcggctgt ctggtctgta 540
actgacgctg aggctcgaaa gcatgggtag cgaacaggat tagataccct ggtagtccat 600
accgtaaacg atgaatgcta agtgttggag ggtttccgcc cttcagtgct gcagctaacg 660
cattaagcat tccgcctggg gagtacgacc gcaaggttga aactcaaagg aattgacggg 720
ggcccgcaca agcggtggag catgtggttt aattcgaagc aacgcgaaga accttaccag 780
gtcttgacat cctttgccca tcgtagagat aaaactttcc cttcggggac aaagtgacag 840
gtggtgcatg gttgtcgtca gctcgtgtcg tgagatgttg ggttaagtcc cgcaacgagc 900
gcaaccctta ttaccagttg ccagcattta gttgggcact ctggtgagac tgccggtgac 960
aaaccggagg aaggtgggga tgacgtcaaa tcatcatgcc ccttatgacc tgggctacac 1020
acgtgctaca atggacggta caacgagttg cgaactcgcg agggtaagct aatctcttaa 1080
agccgatctc agttcggatt gtaggctgca actcgcctac atgaagtcgg aatcgctagt 1140
aatcgcggat cagcacgccg cggtgaatac gttcccgggc cttgtacaca ccgcccgtca 1200
caccatgaga gtttgtaaca cccaaagtcg gtggggtaac ct 1242
<210> 2
<211> 20
<212> DNA
<213> Artificial sequence
<400> 2
agagtttgat cctggctcag 20
<210> 3
<211> 22
<212> DNA
<213> Artificial sequence
<400> 3
tacggytacc ttgttacgac tt 22
<210> 4
<211> 11
<212> PRT
<213> Artificial sequence
<400> 4
Arg Gln Gln Pro Met Thr Leu Asp Tyr Arg Trp
1 5 10
<210> 5
<211> 10
<212> PRT
<213> Artificial sequence
<400> 5
Leu Val Lys Asn Ala Lys Glu Arg Gly Tyr
1 5 10

Claims (3)

1. A lactobacillus corynebacterium for producing broad-spectrum antibacterial peptide is characterized in that: the corynebacterium for producing the broad-spectrum antibacterial peptide is the corynebacterium (L.), (Lactobacillus coryniformis) FZU63, which is classified and named: lactobacillus corynebacterium: (Lactobacillus coryniformis) And is preserved in China general microbiological culture Collection center (CGMCC) in 2020, 4 and 7 months, with the preservation number as follows: CGMCC number 19553, the preservation address is as follows: xilu No.1 Hospital No. 3, Beijing, Chaoyang, North.
2. The broad spectrum antimicrobial peptide-producing Lactobacillus corynebacterium composition AMP-fzu63 of claim 1, wherein: the antibacterial peptide composition AMP-fzu63 comprises 2 polypeptides, wherein the amino acid sequences of the 2 polypeptides are respectively as follows: RQQPMTLDYRW, LVKNAKERGY are provided.
3. The use of the antimicrobial peptide composition AMP-fzu63 of claim 2 for the preservation of marine fish.
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JP2010246398A (en) * 2009-04-10 2010-11-04 Asama Chemical Co Ltd New antibacterial peptide

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JP2010246398A (en) * 2009-04-10 2010-11-04 Asama Chemical Co Ltd New antibacterial peptide

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